The precise interaction of relaxing, contractile and anchoring proteins in brain is now well understood. The objective of this project is to define the regulation of contractile proteins in brain, the anchoring to specific subcellular organelles in brain synaptosomes, specifically as it relates to the uptake, storage and release of neurotransmitters. The coupling in brain of contractile proteins with a relaxing protein complex and the organization and anchoring to specific sites by actin or actin-binding proteins imply an important interaction which has been revealed in part by our reports on the isolation of a complex of relaxing proteins associated with contractile proteins from synaptosomal enriched fractions and an anchoring material isolated from synaptic vesicles, namely alpha-actinin. All the components of this complex system, apparently, are designed to participate in the release of neurotransmitters. In this period, it is intended to investigate if the myosin actin and alpha-actinin molecules in the synaptic vesicles, which are known to bind Mg2 ion ATP, might provide a vehicle for the uptake and storage of Mg2 ion- or Ca2 ion-ATP-biogenic amines, and if the attachment of synaptosomal membrane actin complex to synaptic vesicles is mediated by the anchoring material present on the membranes of these vesicles: alpha-actinin. If such an interaction exists with the association of myosin, energy might be released through the hydrolysis of ATP that will facilitate release of stored, bound neurotransmitters. The regulatory protein complex responds to calcium in a fashion similar to the regulatory proteins of muscle. In brain, however, this apparently permits contractile events to occur between synaptic vesicles and the synaptosomal membranes, accompanied by release of material stored. In this proposal, we intend to study the relaxing proteins, troponin and tropomyosin; the anchoring proteins, alpha-actinin and other actin-binding proteins; and myosin molecules, by determining the biochemical activity; the localization of these proteins within the synaptosomal structures with antibodies that are conjugated with ferritin; and finally, to determine their involvement in the mechanisms of uptake, storage and release of chemical compounds in brain.